Team:RHS-Calgary/Contribution
Contribution
our contribution
The pSBC3-BBa_K2238000 part that codes for our enzyme, produced by Manhattan College iGEM is not included in the distribution kit, so we intend to send the part we create to iGEM so that it can be shared with other teams and utilized in their future endeavours. Let us explain how we plan to do this, and how we are working towards troubleshooting and creating the part. While there were other parts that code for glucose oxidase in the registry, we decided on this particular part that directly corresponded to the specific organism our enzyme is produced by, namely A. niger. With this, we will be using a wild type of GOx, which avoids the potential deletion or inactivation of genes caused by mutations. There was a modified version of the part we chose which included a Cystine-Tag, but we did not go with this part because having an additional tag to the His-Tag already provided in our initial part was unnecessary. Additionally, we were already planning on using a HIS-tagged part due to the materials we intend to use during our protein purification protocol. We already have access to a nickel column, which is capable of purifying specifically HIS-tagged genes. Essentially, the part created by the Manhattan College iGEM team is exactly what we are looking for; it already has a BioBrick Prefix, Suffix, and a TEV Cleavage site; we simply have to recreate it. A TEV-site, which is a part of the BioBrick Prefix, will be useful when removing the Prefix after protein purification. Modifications made to this part by our iGEM team will include, locating sticky ends on both sides of the gene, allowing us to insert it into the standard iGEM plasmid with the corresponding sticky ends.